scholarly journals Moving and stationary actin filaments are involved in spreading of postmitotic PtK2 cells

1993 ◽  
Vol 122 (4) ◽  
pp. 833-843 ◽  
Author(s):  
L Cramer ◽  
TJ Mitchison
Keyword(s):  
Actin Filaments ◽  
Time Lapse ◽  
Forward Movement ◽  
Daughter Cells ◽  
Mechanical Disruption ◽  
Motile Cells ◽  
Postmitotic Cells ◽  
A Cell ◽  
First Time ◽  
Dynamic Populations

We have investigated spreading of postmitotic PtK2 cells and the behavior of actin filaments in this system by time-lapse microscopy and photoactivation of fluorescence. During mitosis PtK2 cells round up and at cytokinesis the daughter cells spread back to regain their interphase morphology. Normal spreading edges are quite homogenous and are not comprised of two distinct areas (lamellae and lamellipodia) as found in moving edges of interphase motile cells. Spreading edges are connected to a network of long, thin, actin-rich fibers called retraction fibers. A role for retraction fibers in spreading was tested by mechanical disruption of fibers ahead of a spreading edge. Spreading is inhibited over the region of disruption, but not over neighboring intact fibers. Using photoactivation of fluorescence to mark actin filaments, we have determined that the majority of actin filaments move forward in spreading edges at the same rate as the edge. As far as we are aware, this is the first time that forward movement of a cell edge has been correlated with forward movement of actin filaments. In contrast, actin filaments in retraction fibers remain stationary with respect to the substrate. Thus there are at least two dynamic populations of actin polymer in spreading postmitotic cells. This is supported by the observation that actin filaments in some spreading edges not only move forward, but also separate into two fractions or broaden with time. A small fraction of postmitotic cells have a spreading edge with a distinct lamellipodium. In these edges, marked actin polymer fluxes backward with respect to substrate. We suggest that forward movement of actin filaments may participate in generating force for spreading in postmitotic cells and perhaps more generally for cell locomotion.

scholarly journals THE TIME OF SYNTHESIS AND THE CONSERVATION OF MITOSIS-RELATED PROTEINS IN CULTURED HUMAN AMNION CELLS

1967 ◽  
Vol 34 (1) ◽  
pp. 97-110 ◽  
Author(s):  
Jesse E. Sisken ◽  
Elaina Wilkes
Keyword(s):  
Time Lapse ◽  
Major Effect ◽  
Human Amnion ◽  
Daughter Cells ◽  
Low Concentrations ◽  
Associated Proteins ◽  
A Cell ◽  
Quantitative Analyses ◽  
Related Proteins ◽  
Kinetics Of

p-Fluorophenylalanine (PFPA), an analogue of phenylalanine which may be incorporated into proteins, increases the duration of mitosis. In the present experiments, based upon quantitative analyses of time-lapse cinemicrographic films, brief treatments of cells with PFPA are shown to affect the duration of metaphase in only those cells which enter division during or shortly after treatment. The offspring of cells with prolonged metaphases also tend to have prolonged metaphases. Analyses of the kinetics of the appearance of prolonged metaphases indicate that some protein specifically associated with mitosis is synthesized primarily during a period which corresponds closely to G2. The manner in which the defect is passed on to daughter cells indicates that the protein involved is conserved and reutilized by daughter cells for their subsequent divisions. Comparable experiments performed with low concentrations of puromycin indicate that the major effect of PFPA is due to its incorporation into protein rather than its ability to inhibit protein synthesis. The fact that puromycin-induced effects can also be passed on to daughter cells is interpreted to mean that cells make only specific amounts of some mitosis-associated proteins and that if a cell "inherits" a deficiency in such protein it is not able to compensate for the deficiency.

scholarly journals Actin Filaments and Myosin I Alpha Cooperate with Microtubules for the Movement of Lysosomes

2001 ◽  
Vol 12 (12) ◽  
pp. 4013-4029 ◽  
Author(s):  
Marie-Neige Cordonnier ◽  
Daniel Dauzonne ◽  
Daniel Louvard ◽  
Evelyne Coudrier
Keyword(s):  
Long Range ◽  
Molecular Motors ◽  
Actin Filaments ◽  
Fluorescent Protein ◽  
Time Lapse ◽  
Myosin I ◽  
Latrunculin A ◽  
Living Mouse ◽  
Green Fluorescent ◽  
First Time

An earlier report suggested that actin and myosin I alpha (MMIα), a myosin associated with endosomes and lysosomes, were involved in the delivery of internalized molecules to lysosomes. To determine whether actin and MMIα were involved in the movement of lysosomes, we analyzed by time-lapse video microscopy the dynamic of lysosomes in living mouse hepatoma cells (BWTG3 cells), producing green fluorescent protein actin or a nonfunctional domain of MMIα. In GFP-actin cells, lysosomes displayed a combination of rapid long-range directional movements dependent on microtubules, short random movements, and pauses, sometimes on actin filaments. We showed that the inhibition of the dynamics of actin filaments by cytochalasin D increased pauses of lysosomes on actin structures, while depolymerization of actin filaments using latrunculin A increased the mobility of lysosomes but impaired the directionality of their long-range movements. The production of a nonfunctional domain of MMIα impaired the intracellular distribution of lysosomes and the directionality of their long-range movements. Altogether, our observations indicate for the first time that both actin filaments and MMIα contribute to the movement of lysosomes in cooperation with microtubules and their associated molecular motors.

scholarly journals Identification of Novel Graded Polarity Actin Filament Bundles in Locomoting Heart Fibroblasts: Implications for the Generation of Motile Force

1997 ◽  
Vol 136 (6) ◽  
pp. 1287-1305 ◽  
Author(s):  
Louise P. Cramer ◽  
Margaret Siebert ◽  
Timothy J. Mitchison
Keyword(s):  
Cell Body ◽  
Actin Filament ◽  
Actin Filaments ◽  
Structural Organization ◽  
Actin Bundle ◽  
Inner Surface ◽  
Filament Bundles ◽  
First Time ◽  
Dynamic Populations ◽  
Uniform Polarity

We have determined the structural organization and dynamic behavior of actin filaments in entire primary locomoting heart fibroblasts by S1 decoration, serial section EM, and photoactivation of fluorescence. As expected, actin filaments in the lamellipodium of these cells have uniform polarity with barbed ends facing forward. In the lamella, cell body, and tail there are two observable types of actin filament organization. A less abundant type is located on the inner surface of the plasma membrane and is composed of short, overlapping actin bundles (0.25–2.5 μm) that repeatedly alternate in polarity from uniform barbed ends forward to uniform pointed ends forward. This type of organization is similar to the organization we show for actin filament bundles (stress fibers) in nonlocomoting cells (PtK2 cells) and to the known organization of muscle sarcomeres. The more abundant type of actin filament organization in locomoting heart fibroblasts is mostly ventrally located and is composed of long, overlapping bundles (average 13 μm, but can reach up to about 30 μm) which span the length of the cell. This more abundant type has a novel graded polarity organization. In each actin bundle, polarity gradually changes along the length of the bundle. Actual actin filament polarity at any given point in the bundle is determined by position in the cell; the closer to the front of the cell the more barbed ends of actin filaments face forward. By photoactivation marking in locomoting heart fibroblasts, as expected in the lamellipodium, actin filaments flow rearward with respect to substrate. In the lamella, all marked and observed actin filaments remain stationary with respect to substrate as the fibroblast locomotes. In the cell body of locomoting fibroblasts there are two dynamic populations of actin filaments: one remains stationary and the other moves forward with respect to substrate at the rate of the cell body. This is the first time that the structural organization and dynamics of actin filaments have been determined in an entire locomoting cell. The organization, dynamics, and relative abundance of graded polarity actin filament bundles have important implications for the generation of motile force during primary heart fibroblast locomotion.

scholarly journals How cortical waves drive fission of motile cells

2020 ◽  
Vol 117 (12) ◽  
pp. 6330-6338 ◽  
Author(s):  
Sven Flemming ◽  
Francesc Font ◽  
Sergio Alonso ◽  
Carsten Beta
Keyword(s):  
Reaction Diffusion ◽  
Cortical Actin ◽  
Animal Cells ◽  
Contractile Ring ◽  
Daughter Cells ◽  
Reaction Diffusion Model ◽  
Motile Cells ◽  
A Cell ◽  
Actin Waves ◽  
Cell Growth And Proliferation

Cytokinesis—the division of a cell into two daughter cells—is a key step in cell growth and proliferation. It typically occurs in synchrony with the cell cycle to ensure that a complete copy of the genetic information is passed on to the next generation of daughter cells. In animal cells, cytokinesis commonly relies on an actomyosin contractile ring that drives equatorial furrowing and separation into the two daughter cells. However, also contractile ring-independent forms of cell division are known that depend on substrate-mediated traction forces. Here, we report evidence of an as yet unknown type of contractile ring-independent cytokinesis that we termed wave-mediated cytofission. It is driven by self-organized cortical actin waves that travel across the ventral membrane of oversized, multinucleatedDictyostelium discoideumcells. Upon collision with the cell border, waves may initiate the formation of protrusions that elongate and eventually pinch off to form separate daughter cells. They are composed of a stable elongated wave segment that is enclosed by a cell membrane and moves in a highly persistent fashion. We rationalize our observations based on a noisy excitable reaction–diffusion model in combination with a dynamic phase field to account for the cell shape and demonstrate that daughter cells emerging from wave-mediated cytofission exhibit a well-controlled size.

Ultrastructure of melanocyte-keratinocyte interactions and pigment transfer in vitro

1978 ◽  
Vol 36 (2) ◽  
pp. 650-651
Author(s):  
Raul I. Garcia ◽  
Evelyn A. Flynn ◽  
George Szabo
Keyword(s):  
Direct Injection ◽  
Skin Pigmentation ◽  
Freeze Fracture ◽  
Pigment Granule ◽  
Time Lapse ◽  
Ultrastructural Level ◽  
Lanthanum Tracer ◽  
A Cell

Skin pigmentation in mammals involves the interaction of epidermal melanocytes and keratinocytes in the structural and functional unit known as the Epidermal Melanin Unit. Melanocytes(M) synthesize melanin within specialized membrane-bound organelles, the melanosome or pigment granule. These are subsequently transferred by way of M dendrites to keratinocytes(K) by a mechanism still to be clearly defined. Three different, though not necessarily mutually exclusive, mechanisms of melanosome transfer have been proposed: cytophagocytosis by K of M dendrite tips containing melanosomes, direct injection of melanosomes into the K cytoplasm through a cell-to-cell pore or communicating channel formed by localized fusion of M and K cell membranes, release of melanosomes into the extracellular space(ECS) by exocytosis followed by K uptake using conventional phagocytosis. Variability in methods of transfer has been noted both in vivo and in vitro and there is evidence in support of each transfer mechanism. We Have previously studied M-K interactions in vitro using time-lapse cinemicrography and in vivo at the ultrastructural level using lanthanum tracer and freeze-fracture.

Modulated polarization microscopy displays the dynamics of cytoskeletal structures in living, motile cells

1995 ◽  
Vol 53 ◽  
pp. 902-903
Author(s):  
J. R. Kuhn ◽  
M. Poenie
Keyword(s):  
Mitotic Spindle ◽  
Actin Filaments ◽  
Cell Shape ◽  
Dynamic Structure ◽  
Living Cells ◽  
Cytoskeletal Proteins ◽  
Polarization Microscopy ◽  
Video Enhancement ◽  
Ultrastructural Studies ◽  
Motile Cells

Cell shape and movement are controlled by elements of the cytoskeleton including actin filaments an microtubules. Unfortunately, it is difficult to visualize the cytoskeleton in living cells and hence follow it dynamics. Immunofluorescence and ultrastructural studies of fixed cells while providing clear images of the cytoskeleton, give only a static picture of this dynamic structure. Microinjection of fluorescently Is beled cytoskeletal proteins has proved useful as a way to follow some cytoskeletal events, but long terry studies are generally limited by the bleaching of fluorophores and presence of unassembled monomers.Polarization microscopy has the potential for visualizing the cytoskeleton. Although at present, it ha mainly been used for visualizing the mitotic spindle. Polarization microscopy is attractive in that it pro vides a way to selectively image structures such as cytoskeletal filaments that are birefringent. By combing ing standard polarization microscopy with video enhancement techniques it has been possible to image single filaments. In this case, however, filament intensity depends on the orientation of the polarizer and analyzer with respect to the specimen.

The locomotory behaviour of small groups of fibroblasts

1965 ◽  
Vol 162 (988) ◽  
pp. 289-302 ◽  
Keyword(s):  
Small Groups ◽  
Control Groups ◽  
Isolated Cells ◽  
Daughter Cells ◽  
A Cell ◽  
Locomotory Behaviour ◽  
Chick Embryo Heart ◽  
Embryo Heart ◽  
Random Nature ◽  
Chemotactic Gradient

Small groups of two to four fibroblasts at the periphery of outgrowths from cultured explants of chick embryo heart were isolated from their neighbours by sweeping away the nearby cells. The groups and the explants were left attached to the glass substrate, undisturbed. The behaviour of the isolated cells was photographically recorded during about 8 h of further culture. The cells of these groups dispersed, though not as a rule so far as to lose all mutual contacts, the dispersal being counterbalanced by the addition of new cells through mitosis. The accompanying changes in speed of locomotion, and the non-random nature of the spreading, are interpreted in terms of the effects of contacts between the cells. During the first four hours after isolation, but not thereafter, the cells of the groups on the average moved slowly away from the explant. Control groups in an intact outgrowth moved away faster and with no diminution of speed during the period of observation. The movement of the isolated groups can be partly accounted for by the tendency of cells to conserve for a time the direction of their movement before isolation; and by a strong reluctance of the isolated cells to move across the area, from which cells had been scraped away, that lay between the group and the explant. A new outgrowth of the residual sheet of cells still connected to the explant, however, advanced across this area, approaching and in most cases overhauling the isolated group. It is concluded that a chemotactic gradient around the explant is unlikely to play any significant part in the outward movement of fibroblasts from an explant in tissue culture. The cells of the isolated groups underwent an outburst of mitosis about 3 h after isolation. Mitoses in these relatively free cells are oriented in relation to the polarity of the cell before division. Locomotion of the daughter-cells tends to be faster than usual for at least 2 h after a cell divides.

scholarly journals Characterization of a Novel Fructosyltransferase from Lactobacillus reuteri That Synthesizes High-Molecular-Weight Inulin and Inulin Oligosaccharides

2002 ◽  
Vol 68 (9) ◽  
pp. 4390-4398 ◽  
Author(s):  
S. A. F. T. van Hijum ◽  
G. H. van Geel-Schutten ◽  
H. Rahaoui ◽  
M. J. E. C. van der Maarel ◽  
L. Dijkhuizen
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Lactobacillus Reuteri ◽  
Bacterial Strains ◽  
Potential Health ◽  
Isolation And Characterization ◽  
A Cell ◽  
Encoding Gene ◽  
First Time

ABSTRACT Fructosyltransferase (FTF) enzymes produce fructose polymers (fructans) from sucrose. Here, we report the isolation and characterization of an FTF-encoding gene from Lactobacillus reuteri strain 121. A C-terminally truncated version of the ftf gene was successfully expressed in Escherichia coli. When incubated with sucrose, the purified recombinant FTF enzyme produced large amounts of fructo-oligosaccharides (FOS) with β-(2→1)-linked fructosyl units, plus a high-molecular-weight fructan polymer (>107) with β-(2→1) linkages (an inulin). FOS, but not inulin, was found in supernatants of L. reuteri strain 121 cultures grown on medium containing sucrose. Bacterial inulin production has been reported for only Streptococcus mutans strains. FOS production has been reported for a few bacterial strains. This paper reports the first-time isolation and molecular characterization of (i) a Lactobacillus ftf gene, (ii) an inulosucrase associated with a generally regarded as safe bacterium, (iii) an FTF enzyme synthesizing both a high molecular weight inulin and FOS, and (iv) an FTF protein containing a cell wall-anchoring LPXTG motif. The biological relevance and potential health benefits of an inulosucrase associated with an L. reuteri strain remain to be established.

scholarly journals Contribution of Filopodia to Cell Migration: A Mechanical Link between Protrusion and Contraction

2010 ◽  
Vol 2010 ◽  
pp. 1-13 ◽  
Author(s):  
Fei Xue ◽  
Deanna M. Janzen ◽  
David A. Knecht
Keyword(s):  
Cell Migration ◽  
Actin Polymerization ◽  
Myosin Ii ◽  
Temporal Distribution ◽  
Leading Edge ◽  
Distal Portion ◽  
Actin Binding ◽  
Actin Networks ◽  
Motile Cells ◽  
A Cell

Numerous F-actin containing structures are involved in regulating protrusion of membrane at the leading edge of motile cells. We have investigated the structure and dynamics of filopodia as they relate to events at the leading edge and the function of the trailing actin networks. We have found that although filopodia contain parallel bundles of actin, they contain a surprisingly nonuniform spatial and temporal distribution of actin binding proteins. Along the length of the actin filaments in a single filopodium, the most distal portion contains primarily T-plastin, while the proximal portion is primarily bound byα-actinin and coronin. Some filopodia are stationary, but lateral filopodia move with respect to the leading edge. They appear to form a mechanical link between the actin polymerization network at the front of the cell and the myosin motor activity in the cell body. The direction of lateral filopodial movement is associated with the direction of cell migration. When lateral filopodia initiate from and move toward only one side of a cell, the cell will turn opposite to the direction of filopodial flow. Therefore, this filopodia-myosin II system allows actin polymerization driven protrusion forces and myosin II mediated contractile force to be mechanically coordinated.

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